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Protocols to detect senescence-associated beta-galactosidase (SA-βgal) activity, a biomarker of senescent cells in culture and in vivo

Nature Protocols volume 4, pages 1798–1806 (2009)Cite this article

Abstract

Normal cells can permanently lose the ability to proliferate when challenged by potentially oncogenic stress, a process termed cellular senescence. Senescence-associated beta-galactosidase (SA-βgal) activity, detectable at pH 6.0, permits the identification of senescent cells in culture and mammalian tissues. Here we describe first a cytochemical protocol suitable for the histochemical detection of individual senescent cells both in culture and tissue biopsies. The second method is based on the alkalinization of lysosomes, followed by the use of 5-dodecanoylaminofluorescein di-β-D-galactopyranoside (C12FDG), a fluorogenic substrate for βgal activity. The cytochemical method takes about 30 min to execute, and several hours to a day to develop and score. The fluorescence methods take between 4 and 8 h to execute and can be scored in a single day. The cytochemical method is applicable to tissue sections and requires simple reagents and equipment. The fluorescence-based methods have the advantages of being more quantitative and sensitive.

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Figure 1: Senescence-associated beta-galactosidase (SA-βgal) detection in replicative senescence and stress-induced premature senescence.

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Acknowledgements

O.T. and F.D.-C. are respectively Senior Research Associate and Research Associate of the FNRS, Belgium. We acknowledge the Région Wallonne for the First-DEI 'Cosmet-X' project, the Network II Senegene project and the Pole of Excellence Nanotoxico project. We thank the European Commission for the Integrated Projects Proteomage (LSHM-CT-2005-518230) and GeHA (LSHM-CT-2004-503270); for the Coordination & Support Actions Link-Age and WhyWeAge (LSHM-CT-2005-513866, HEALTH-F4-2008-200970); and the Marie Curie Matiss Project (MTKI-CT-2006–042768). Also supported by grants from the US National Institutes of Health (AG09909, AG017242, CA0126540, AG032117 and AG025708) and the British Heart Foundation.

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Authors and Affiliations

  1. Department of Biology, Faculty of Sciences, Research Unit on Cellular Biology (URBC), University of Namur (FUNDP), Namur, Belgium

    Florence Debacq-Chainiaux & Olivier Toussaint

  2. Centre for Biomedical Research, Cardiff School of Health Sciences, University of Wales Institute, Cardiff (UWIC), Cardiff, UK

    Jorge D Erusalimsky

  3. Buck Institute for Age Research, Novato, California, USA

    Judith Campisi

  4. Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California, USA

    Judith Campisi

Authors
  1. Florence Debacq-Chainiaux
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  2. Jorge D Erusalimsky
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Contributions

F.D.-C., O.T., J.D.E. and J.C. carried out and analyzed experiments. F.D.-C., O.T., J.D.E. and J.C. wrote the paper.

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Correspondence to Olivier Toussaint.

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Debacq-Chainiaux, F., Erusalimsky, J., Campisi, J. et al. Protocols to detect senescence-associated beta-galactosidase (SA-βgal) activity, a biomarker of senescent cells in culture and in vivo. Nat Protoc 4, 1798–1806 (2009). https://doi.org/10.1038/nprot.2009.191

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